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Protein absolute quantification (AQUA) uses stable isotope labeled synthetic peptides to quantify the amount of a corresponding native protein in a sample. In protein AQUA, a synthetic peptide nearly identical to the protein to be quantified is generated. The only difference is the incorporation of one stable isotope labeled amino acid. The protein sample to be quantified is mixed with a known amount of AQUA peptide and protease digested. The digested peptides are separated by chromatography and then analyzed by mass spectrometry (MS). Since the AQUA peptide is a replica of the native protein, except slightly heavier, the two peptides elute, migrate, and ionize together but are distinguishable by MS. Comparison of the native protein to the known amount of AQUA peptide allows absolute quantification of the native protein. Protein AQUA eliminates the labeling of native proteins and works for low abundance proteins. Protein AQUA can be used to evaluate post-translational modifications of proteins and for clinical biomarker testing. Specific clinical applications are testing for staphlococcal toxic shock syndrome and monitoring patient response to chemotherapy. (Credit: Brooke Anderson-White)
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